Accelerated graft arteriosclerOsis (AGA) represents a major obstacle to long term survival of heart transplant recipients. While it has become clear that AGA is a multifactorial problem, the specific mechanisms leading to AGA remain unknown. Because of their pivotal role in the maintenance of vessel homeostasis, endothelial cells (EcS) are likely to represent prime targets in the pathogenesis of AGA. Project 1 is designed to provide in-depth characterization of the process that affects ECs in AGA, from extracellular factors to intracellular molecular pathways that transduce messages to the genome of ECs. By integrating our studies on ECs with other projects of this program, we intend to provide information that will be key to the development of specific strategies to suppress AGA. In AGA, the functional damage to the endothelium is diffuse. Morphologically, areas of endothelial erosion can be found dispersed along the coronary vessels. Several mechanisms of BC ii-jury have been implicated in AGA: (i) oxidative injury resulting from ischemia/reperfusion at the time of surgery; (ii) immunological injuries involving complement activation, processing and presentation of antigens, activation and adhesion of T.lymphocytes, in particular cytolytic T- lymphocytes (CTL), sequestration of monocytes, production of antibodies against allo- or iso-antigens, and dysregulation of the immune system with Cyclosporine A; (iii) injury with CMV-infection/reactivation; and (iv) direct Cyclosporine A toxicity. These various mechanisms could contribute either independently or in concert, to the diffuse abnormalities of ECs in AGA. Particular emphasis will be placed on the following pathway, which will provide the basis for the specific aims of this project: (i) the initial injury results in uncoupling of Gi-2 from its membrane receptors; (ii) the uncoupling of Gi-2 then results in BC dysfunction and overexpression of Fas on the surface of ECs, through a pathway that requires NF-kappaB activation; (iii) overexpression of Fas, in turn, results in the "kiss of death" as CTL bind to the ECs through the interaction of Fas and Fas-ligand present on the CTL surface; (iv) increased BC apoptosis will further aggravate the dysfunctional aspect of the endothelium and promote fibrin production and deposition due to the abnormal presence of phosphatidylserine on the surface of ECs. As a result, the endothelium in these patients is not capable of carrying out its usual functions namely, vasodilation, anti-thrombotic effect, anti- proliferative effect for the cells of the vessel wall, control of the attachment of inflammatory cells, and homeostasis of extracellular protein production, leading to the development of AGA.